Pellicle Stimulon in Mycobacterium tuberculosis
Ontology highlight
ABSTRACT:
ORGANISM(S): Mycobacterium tuberculosis
SUBMITTER: Brian Weinrick
PROVIDER: E-GEOD-37392 | biostudies-arrayexpress |
REPOSITORIES: biostudies-arrayexpress
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Project description:Triplicate 10mL cultures of M. tuberculosis grown in Sautons media without detergent in planktonic (exponential growth phase at OD600 0.2) compared to pellicle (3 weeks growth without shaking in 5% CO2) conditions.
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2013-05-24 | E-GEOD-42151 | biostudies-arrayexpress
Project description:New chemotherapeutics are urgently required to control the tuberculosis pandemic fueled by the emergence of multidrug- and extensively-drug-resistant Mycobacterium tuberculosis strains and the bacterium`s catastrophic alliance with HIV. Here we describe a novel trehalose-to-α-glucan pathway in M. tuberculosis comprising four enzymatic steps mediated by TreS, Pep2, GlgB, and GlgE, identified as an essential maltosyltransferase capable of utilizing maltose 1-phosphate. Using traditional and chemical reverse genetics, we show that GlgE inactivation causes rapid death of M. tuberculosis in vitro and in mice, through self-poisoning by maltose 1-phosphate accumulation driven by a self-amplifying feedback loop promoting pleiotropic phosphosugar-induced stress responses. Moreover, this α-glucan pathway exhibited a synthetic lethal interaction with the glucosyltransferase Rv3032 involved in biosynthesis of specialized α-glucan derivatives. The unique combination of gene essentiality within a synthetic lethal pathway validates GlgE as a new class of drug targets, revealing novel synergistic mechanisms to induce death in M. tuberculosis. Transcriptional profiling was performed to characterize the lethality induced by maltose 1-phosphate accumulation. Triplicate 10 mL cultures of the conditional lethal Mtb mutant strain H37Rv Delta treS Delta glgE (pMV361::treS) and of the vector control strain H37Rv Delta treS Delta glgE (pMV361) were grown in liquid culture to log-phase in the presence of 5 mM validamycin A (VA) to suppress M1P formation. Subsequently, cells were washed to remove the inhibitor; after 48 h of starvation for VA cultures were harvested. Keywords: tuberculosis, trehalose, compound treatment design, genetic modification design, and stimulus or stress design Three biological replicates with one dye-flip
2010-05-14 | E-GEOD-18575 | biostudies-arrayexpress
Project description:The emergence of drug resistance among tuberculosis (TB) patients is often associated with their non-compliance to the length of the chemotherapy, which can reach up to 2 years for the treatment of multi-drug-resistant (MDR) TB. Drugs that would kill TB faster and would not lead to the development of drug resistance could shorten chemotherapy significantly. In Escherichia coli, the common mechanism of cell death by bactericidal antibiotics is the generation of highly reactive hydroxyl radicals via the Fenton reaction. Since ascorbic acid (vitamin C) is known to drive the Fenton reaction, we tested whether the Fenton reaction could lead to a bactericidal event in Mycobacterium tuberculosis by treating M. tuberculosis cultures with vitamin C. Here, we report that the addition of vitamin C to drug-susceptible, MDR and extensively drug-resistant (XDR) M. tuberculosis strains results in sterilization of the cultures in vitro. We show that the sterilizing effect of vitamin C on M. tuberculosis was dependent on the production of high ferrous ion levels and reactive oxygen species. Although, this potent sterilizing activity of vitamin C against M. tuberculosis in vitro was not observed in mice, we believe this activity needs further investigation. Comparison of vitamin C treated Mycobacterium tuberculosis transcriptome relative to untreated; Three biological replicates, second is a dye flip
2013-05-24 | E-GEOD-42293 | biostudies-arrayexpress
Project description:Transcriptional profiling of Mycobacterium tuberculosis H37Rv grown in minimal media with or without detergent for 5 days to investigate basis of differences in vaccine efficacy dependent on growth conditions Two condition experiment, detergent grown vs. grown without detergent. Biological replicates: 3 of each condition, independently grown and harvested. One replicate per array, third array is a dye-flip.
2016-05-31 | E-GEOD-72745 | biostudies-arrayexpress
Project description:Mycobacteria can synthesize NAD+ using either the de novo biosynthesis pathway or the salvage pathway. The deletion of the three genes involved specifically in the NAD+ de novo biosynthesis pathway in the human pathogen Mycobacterium tuberculosis had no effect on the growth of the strain in vivo. In contrast, the same deletion in the bovine pathogen Mycobacterium bovis resulted in a strain that could not grow in vivo and could only grow in vitro with substantial nicotinamide supplmentation. This striking difference was attributed to the known defect in the nicotinamidase PncA of M. bovis, since introducing the M. tuberculosis pncA gene into the M. bovis strain defective for de novo NAD+ biosynthesis restored growth in vitro and in vivo. This study demonstrates that NAD+ starvation is a cidal event in mycobacteria and confirms that enzymes common to the de novo and salvage pathways may be good drug targets. We also propose that simultaneously targeting both the salvage and the de novo NAD+ biosynthesis pathways represents a potentially effective way to treat infection with tubercle bacilli. To characterize the lethality induced by nicotinamide starvation transcriptional profiling of the auxotrophs was performed. Triplicate 50 mL cultures of M. tuberculosis and M. bovis Delta nadABC mutants were grown in 7H9 OADC glycerol 0.05% tween broth in 500 mL roller bottles to an OD600nm= 0.1 in a roller incubator at 37°C. The cells were washed 1x in PBS and resuspended in 50 mL 7H9 OADC glycerol 0.05% tween broth with or without 20mg/L nicotinamide and returned to the incubator. After 7 days, cultures were harvested. Three biological replicates of each of two species with one dye-flip each
2010-05-14 | E-GEOD-18909 | biostudies-arrayexpress
Project description:Mechanisms governing Mycobacterium tuberculosis acid-fastness and its capacity to induce long-term infections remain unknown. Serine/Threonine phosphorylation represents an emerging theme allowing mycobacteria to adapt their cell envelope structure/composition in response to environmental changes. We addressed whether phosphorylation of KasB, a mycolic acid biosynthetic enzyme, modulates M. tuberculosis pathogenicity. Phosphorylation of KasB occurred at Thr334 and Thr336 in vitro and in mycobacteria. A mutant strain bearing an kasB_T334D/T336D allele, mimicking constitutive KasB phosphorylation, was generated by specialized linkage transduction. This resulted in shortened mycolic acids and the lack of trans-cyclopropanation. Structural/modeling analyses revealed Thr334 and Thr336 in the vicinity of the catalytic triad, implying that phosphorylation of these residues impaired KasB activity. Importantly, the phosphomimetic strain lost acid-fast staining and was more attenuated than a kasB deletion mutant in immunocompetent and immunodeficient mice. The absence of lung pathology and mortality infers this mutant to represent a valuable vaccine candidate. This work emphasizes the critical role of Ser/Thr kinase-dependent signaling in controlling mycolic acid elongation, acid-fastness, virulence and has important clinical implications for diagnosis of latent infections. Transcriptome of kasB null, phosphoablative, and phosphomimetic mutants compared to parental. Triplicate 10ml cultures of M. tuberculosis CDC1551 and kasB null, phosphoablative, and phosphomimetic mutants were grown to OD 1.0 and harvested for transcriptional profiling.
2014-04-01 | E-GEOD-47640 | biostudies-arrayexpress
Project description:We compared genetic profiles of planktonic stage to biofilm stage of deep sea bacterium Pseudoalteromonas sp. SM9913 and revealed genetic features during switch from planktonic to pellicle stage in Pseudoalteromonas sp. SM9913. mRNA profiles of Pseudoalteromonas sp. SM9913 planktonic cells, initial pellicle cells and mature pellicle cells were generated by Illumina Hiseq2000.
2015-11-03 | E-GEOD-74569 | biostudies-arrayexpress
Project description:To investigate the gene expression profile of pellicle cells of Pseudomonas aeruginosa, microarray analysis was performed. Transcriptome profiles of pellicle cells and planktonic cells grown in LB medium were determined by Affymetrix GeneChip. Gene expression pattern that is specific to pellicle cells was evaluated by comparing the data set with that of planktonic cells.
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Project description:Shewanella oneidensis is an important model organism for bioremediation studies because of its diverse respiratory capabilities. In recent years, biofilm development of S. oneidensis has been extensively studied because it is essential to reduce solid metals. As a special form of biofilm, however, pellicles are largely overlooked. The goal of this work was to understand requirements of S. oneidensis pellicle formation and the molecular basis of pellicle formation. We demonstrated that successful pellicle formation and survival was likely to require the threshold level of cell density and higher concentration of oxygen. Proteinase K and EDTA were potent pellicle disrupter. DNA microarray experiments were used to study the gene expression profile of young air–liquid interface pellicle relative to planktonic cells, which indicated that the air–liquid interface pellicle was more metabolically active than the planktonic cells. Most notably, consistently up-regulation of iron or heme uptake and transportation proteins was observed in the S. oneidensis MR-1 pellicle. However, neither the hmuT nor hugA heme transport mutant was defective in pellicle formation. An examination of the influence of several metal cations on the anti-pellicle activity of EDTA showed that Ca (II), Mn(II), Cu(II), and Zn(II) fully protected S. oneidensis MR-1 pellicle against EDTA treatment while additional of iron enabled the initiation of pellicle formation but maturation was significantly impaired. Collectively, iron was less important than other metals with respect to pellicle formation in S. oneidensis. A fresh colony grown overnight on a LB plate was used to inoculate 50 ml LB and incubated in a shaker (200 rpm) to an OD600 of 0.8 at the room temperature. This culture was then diluted 500-fold with fresh LB, resulting in the starting cultures. Aliquots of 30ml starting cultures were transferred to 50-ml Pyrex beakers and allowed to develop pellicles at the room temperature. When a complete but thin (young pellicle) at the interface were formed (about 30h hours), planktonic culture and pellicle were separated and applied to centrifugation at 8000 rpm for 3 min at room temperature. 3 parallel starting cultures were used and 3 samlpes of pellicle cells or planktonic cells were collected at 30h. RNA from the pellicle cells was fluorescently labeled with Cy3, and that from the planktonic was labeled with Cy5.
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